Redox regulation of the NMDA channel has been demonstrated for primary neurons and recombinant proteins transfected into cells. Reduction of the channel increases whole cell currents while oxidation reduces currents. Site directed mutagenesis demonstrated the presence of critical cysteine groups on the NMDA channel proteins. Our recent data confirms the regulation of NMDA channel function by redox factors and further demonstrates that nitric oxide (NO) alters NMDA channel function increasing whole cell currents by increasing channel open probability. Since the NMDA channel is involved in a putative cellular form of learning and memory, long term potentiation (LTP), and has also been implicated in the excitotoxicity associated with various neuropathological disease states, alteration of the NMDA channel by the local tissue redox state may be of significance to the functional changes associated with these processes. The local tissue redox state, in turn, may be regulated by the generation of reactive oxygen species (including NO) into the environment of the NMDA channel. Microglia, the CNS macrophage, produce superoxide anion when stimulated as part of the respiratory burst and NO is produced when the microglia are stimulated with the appropriate cytoactive factors. Microglia also have cytoplasmic processes which abut synaptic endings and are involved in synaptic stripping. Thus, microglia are in the appropriate location to alter NMDA channel function by the production of reactive oxygen species. We will test the hypothesis that microglia alter NMDA channel function by using a sniffing patch technique. Outside-out patches will be prepared from HEK cells transfected with recombinant NMDA channel proteins. These patches will then be used to determine the effect of microglial-produced superoxide anion or NO on NMDA single channel parameters. Changes in single channel conductance, open probability, open duration and burst duration will be assessed in untreated microglia and microglia that have been induced to produce either superoxide anion or NO. The microglia will then be pretreated with lipopolysaccharide, (LPS), an inflammatory mediator, with interferon, an immunological mediator; or with Alphabeta peptide, a a microglial activator relevant to Alzheimer s disease. These factors are known to increase the activation of the microglia, promoting their cytotoxicity. Specificity of the effect will be determined using known antioxidants and a cell line engineered to generate NO.